Sasha E. Stanton

Variation in the Incidence and Magnitude of Tumor-Infiltrating Lymphocytes in Breast Cancer Subtypes: A Systematic Review

Importance The presence of tumor-infiltrating lymphocytes (TILs) is a favorable prognostic factor in breast cancer, and TILs may synergize with chemotherapy and immune checkpoint inhibitor therapy for improved clinical response. A more detailed understanding of the variation in lymphocytic infiltration in breast cancer may aid in identifying subtypes more amenable to immunomodulation. Objective To determine the median percentage of patients with breast cancer with no, intermediate, or high levels of TIL and assess variations in lymphocytic cell subsets across breast cancer subtypes. Evidence Review Eligible studies (PubMed, 1990-2015) analyzed tumor lymphocytic, CD8+, and FOXP3+ cellular infiltrates, and used multivariable analyses and quantitative methods for enumerating cell populations. Selection of of studies was performed in accordance with PRISMA guidelines and evaluated by 2 independent appraisers. Findings Fifteen studies (n = 13 914) met prespecified criteria and were reviewed in December 2015. A median of 11% (range, 5%-26%) of breast cancers demonstrate lymphocyte-predominant breast cancer (LPBC), with approximately 16% of cancers showing no evidence of TILs. Triple-negative (TN) breast cancers demonstrated the highest incidence of LPBC (20%; range, 4%-37%). This incidence is similar to that of breast cancers that are human epidermal growth factor 2 positive and either hormone receptor positive or negative (HER2+) at 16% (range 11%-24%). Hormone receptor positive/HER2- (HR+) breast cancer showed the lowest incidence of LPBC at 6% (range, 3%-12%). CD8+ T-cell infiltrates, indicative of type I immunity, were found in 48% of all breast cancers (range, 32%-80%) with similar levels observed in TN (60%; range, 40%-91%) and HER2+ disease (61%; range, 40%-83%). Fewer HR+ tumors demonstrated CD8+ TIL (43%; range, 30%-73%). The highest levels of FOXP3+ cells were observed in TN (70%; range, 65%-76%) and HER2+ disease (67%; range, 61%-74%). A minority of HR+ breast cancers demonstrated high levels of tumor-infiltrating FOXP3+ cells (38%; range, 35%-41%). Conclusions and Relevance The magnitude of TIL is variable within and between breast cancer subtypes. Levels of lymphocytic subpopulations may identify breast cancers more amenable to immunomodulation and indicate additional strategies to enhance immunity in patients with low to moderate levels of TILs.

Triple-negative breast cancer: immune modulation as the new treatment paradigm.

Recent studies of tumor lymphocytic immune infiltrates in breast cancer have suggested an improved prognosis associated with increasing levels of tumor-infiltrating lymphocytes (TIL). Triple-negative breast cancer (TNBC) is the breast cancer subtype that has the greatest incidence of patients with a robust tumor immune infiltrate, although it is still a minority of patients. Elevated levels of either intratumoral or stromal T cells are associated with an improved overall survival (OS) and disease-free survival (DFS) in TNBC as compared with other breast cancer subtypes. TNBC may be immunogenic for several reasons. Subtypes of TNBC have a significant number of genetic mutations, and the immune system may see the aberrant proteins encoded by these mutations as foreign. Moreover, TNBC is associated with a prognostic gene signature that also includes B cells. Antibodies secreted by B cells may bind to tumor antigens and amplify the adaptive immune response that has already been initiated in the tumor. New immune modulatory agents, including immune checkpoint inhibitors, have shown activity in immunogenic tumors such as melanoma and bladder cancer and have recently been tested in TNBC. The clinical response rates observed, patterns of response, and adverse event profiles are similar to what has been described in melanoma where this class of agents has already been approved for clinical use in some cases. Lessons learned in assessing the immunogenicity of TNBC, potential mechanisms of immune stimulation, and response to immune modulatory drugs lay the foundation for the development of immune-based therapies in all subtypes of the disease.

Can Immunity to Breast Cancer Eliminate Residual Micrometastases

An effective immune response has the potential for breast cancer sterilization with marked reduction in the potential for disease relapse. Adaptive type I immune cells uniquely have the capability of (i) cytotoxic T-cell activation and proliferation until all antigen expressing cells are eradicated, (ii) traversing endothelial barriers to penetrate tumor deposits wherever they occur, and (iii) immunologic memory, which allows the persistence of destructive immunity over the years it may take for breast cancer micrometastases to become clinically evident. Numerous recent investigations suggest that some breast cancers stimulate the type of immunity that results in a decreased risk of relapse. Moreover, the endogenous type I tumor microenvironment or type I immunity induced by drugs or biologic agents may improve response to standard therapies, further lowering the probability of disease recurrence. Clin Cancer Res; 19(23); 6398–403. ©2013 AACR.

The Antigenic Repertoire of Premalignant and High-Risk Lesions

Prophylactic vaccines have been a major advance in preventing the development of infections after exposure to pathogens. When contemplating an effective approach to cancer prevention, vaccines offer unique advantages over other more standard approaches: First, once appropriately stimulated, antigen-specific T cells will travel to all sites of disease and eradicate cells bearing the proteins to which the T cells have been primed by vaccination. Second, successful immunization will further result in the development of immunologic memory, providing lifelong immunologic surveillance. There is evidence of an adaptive tumor immune infiltrate even at the earliest stages of breast and colon cancer development. Furthermore, there is measurable immunity to lesion-associated antigens present in patients who will eventually develop malignancy even before cancer is clinically evident. Recent studies are beginning to unmask the preinvasive antigenic repertoire for these two malignancies. Preliminary experiments in transgenic mouse models of mammary and intestinal tumors suggest that immunization against antigens expressed in preinvasive and high-risk lesions may be effective in preventing the development of invasive malignancy. Cancer Prev Res; 8(4); 266–70. ©2015 AACR.

Current methods of epitope identification for cancer vaccine design

The importance of the immune system in tumor development and progression has been emerging in many cancers. Previous cancer vaccines have not shown long-term clinical benefit possibly because were not designed to avoid eliciting regulatory T-cell responses that inhibit the anti-tumor immune response. This review will examine different methods of identifying epitopes derived from tumor associated antigens suitable for immunization and the steps used to design and validate peptide epitopes to improve efficacy of anti-tumor peptide-based vaccines. Focusing on in silico prediction algorithms, we survey the advantages and disadvantages of current cancer vaccine prediction tools.

Immunotherapy in breast cancer: An introduction

The field of breast cancer immunology has progressed tremendously over the last decade. Twenty years ago immunotherapy was not considered for the treatment of breast cancers because breast cancer was not considered immunogenic. Today we know that most patients with breast cancer have some evidence of an adaptive immune response against their tumors, detectable either in the peripheral blood or in the tumor. Moreover, immunity to breast cancer begins at the earliest stages of the disease, in some patients prior to diagnosis. Recent evidence suggests that lymphocytes infiltrating breast cancers and found in the tumor stroma are strong prognostic indicators of a beneficial disease outcome. These observations now pave the way for the integration of immunomodulation into standard of care therapy for the treatment of breast cancer.

Designing vaccines to prevent breast cancer recurrence or invasive disease.

With the emerging importance of the tumor immune environment in both invasive and preinvasive breast cancer, vaccine therapy may provide a well-tolerated, durable therapy for preinvasive breast cancer by preventing both recurrent preinvasive disease and progression to invasive disease. Preinvasive breast cancer is ideal for vaccines because the disease is slow growing and the patients are not immunosuppressed. However, design of the vaccines are critical including consideration of appropriate antigens, selecting Th1 stimulating epitopes, and using multiantigen vaccines to address tumor heterogeneity.

Immunotherapy for breast cancer: is it feasible?

Historically breast cancer has been considered immunologically silent. Patients have had limited access to the types of immunotherapy available to melanoma and lung cancer patients, but this could all be set to change as recent preclinical and clinical studies have highlighted the potential of immunotherapy for breast cancer. Breast cancer is now one of the major cancer types for which new immune-based treatments are being developed. Immunotherapy for breast cancer: is it feasible?

Immunologic Approaches to Breast Cancer Therapy

Although breast cancer tumors tend to have less tumor immune infiltration than immunogenic tumors such as melanoma and colon cancer, the prognostic importance of tumor immune infiltrates in breast cancer has confirmed the essential role of the immune response to the breast tumor. Increased tumor immune infiltrate, particularly antitumor type 1 (Th1) immune infiltrate, has been shown to predict improved disease-free and overall survival, particularly in triple-negative breast cancer, and to influence the response to chemotherapy. Furthermore, autoantibodies against tumor antigens are present in patients even before breast cancer development but not in women who will not develop breast cancer, and these autoantibodies may provide both early diagnosis and response to therapy biomarkers. Finally, the generation of clinical responses to emerging therapies including vaccines and immune checkpoint inhibitor monoclonal antibodies has clearly demonstrated that breast cancer is immunologically active. In the future, therapies that harness the immune system will be used in all stages of breast cancer treatment including the possibility of stimulating immunity for breast cancer prevention. This chapter provides an overview of current immunologic approaches to develop predictive and prognostic assays in breast cancer and novel, clinically effective immune therapy; it also provides an overview of considerations for new directions in breast immunooncology for the future.

Abstract 2224: Panel of autoantibodies to seven early tumor-associated overexpressed proteins can identify women with DCIS from women with benign breast tumors detected by mammography

While mammography is essential to identify early breast cancer, the sensitivity is dependent on breast characteristics. Currently all breast masses identified by mammography have to be biopsied to determine if malignant. A serum-based biomarker used along with mammography to identify pre-malignant lesions to biopsy would both improve the sensitivity of mammography and reduce the number of biopsies needed. We have identified a panel of autoantibodies found in early breast cancer. An autoantibody biomarker is ideal because antibody immunity can be detected with very low levels of antigen with direct antigen recognition by B cells resulting in clonal amplification of antigen specific plasma cells. Work in our laboratory has identified tumor-associated proteins present in pre-malignant tumors that are necessary for survival of human breast cancer cells across breast cancer subtypes. We have found that increased autoantibodies to seven of these early tumor-associated proteins (PDIA6, KRT8, SERBP1, ARPC2, RRM2, AURKA, and NDC80) were present in the sera of women with pre-malignant breast atypia but not women with no breast atypia or benign breast atypia and may be a panel to increase sensitivity of mammography. The presence of autoantibodies was evaluated in 191 individuals, 36 women with no breast atypia, 12 women with benign breast atypia, 36 patients with fibroadenoma, 12 patients with hyperplasia, 59 patients with ductal carcinoma in situ (DCIS), and 36 patients with invasive breast cancer (IBC). We found more women with pre-malignant breast atypia had increased autoantibodies to at least one of the seven early tumor associated proteins as compared to women with either no breast atypia or benign breast atypia. For example, a positive autoantibody response is over 2 standard deviations above the mean found in women with no breast atypia. There were 2.9% of women with positive autoantibody response to RRM2 in women without breast atypia. However, in women with breast atypia, there was a positive antibody response in 41.7% of individuals with hyperplasia, 48.6% of individuals with fibroadenoma, 34.5% of individuals with DCIS, and 28.6% of individuals with IBC. All seven autoantibodies could predict patients with hyperplasia, fibroadenoma, DCIS, and IBC from both women with no atypia and women with benign breast atypia. For example, the seven-antibody panel could identify DCIS from women with no breast atypia with AUC of 0.95 (95% CI 0.912 to 0.995) and from women with benign breast atypia with AUC of 0.71 (95% CI 0.519 to 0.898). This is a better AUC than seen with previous published autoantibody profiles in DCIS. Future studies will validate these findings with the goal of developing a serum biomarker that will improve sensitivity of mammography. Citation Format: Sasha Elizabeth Stanton, Erik Ramos, Mary L. Disis. Panel of autoantibodies to seven early tumor-associated overexpressed proteins can identify women with DCIS from women with benign breast tumors detected by mammography [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2224.